Relationship between the shape and the membrane potential of human red blood cells

1984 ◽  
Vol 82 (1) ◽  
pp. 1-13 ◽  
Author(s):  
Ellen M. Bifano ◽  
Terri S. Novak ◽  
Jeffrey C. Freedman
Keyword(s):  
Membrane Potential ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Human Red Blood Cells

Proton-sulfate cotransport: external proton activation of sulfate influx into human red blood cells

1984 ◽  
Vol 247 (3) ◽  
pp. C247-C259 ◽  
Author(s):  
M. A. Milanick ◽  
R. B. Gunn
Keyword(s):  
Membrane Potential ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Half Cycle ◽  
Ph Values ◽  
Human Red Blood Cells ◽  
Internal Ph ◽  
Ph Range ◽  
External Ph ◽  
Chloride Concentrations

Sulfate influx into human red blood cells was measured at 0 and 22 degrees C at several fixed external pH values between 3 and 10. These cells had normal internal pH and chloride concentrations so that sulfate influx was not limited by the efflux half-cycle reactions. The flux was a Michaelis-Menten function of sulfate concentration at each pH with K1/2SO4 = 4-10 mM. External protons activated influx 100-fold at a single site with a pK = 5.9 at 22 degrees C and 5.5 at 0 degrees C. This pK is similar to the value 5.99 +/- 0.3 for external proton binding to the sulfate-loaded transporter at 0 degrees C (J. Gen. Physiol. 79: 87-114, 1982). The flux was stilbene sensitive even in valinomycin-treated cells and was independent of membrane potential. This proton-activated influx appears to be proton-sulfate cotransport. At high pH there was a proton-independent flux that was membrane potential and stilbene sensitive. This proton-insensitive flux appears to be SO4(2-)/Cl- exchange or net sulfate influx. The sulfate influx over the entire pH range may be described in terms of an equation for the sum of the influxes through these two pathways on band 3.

scholarly journals Proton (or hydroxide) fluxes and the biphasic osmotic response of human red blood cells.

1993 ◽  
Vol 102 (1) ◽  
pp. 99-123 ◽  
Author(s):  
J D Bisognano ◽  
J A Dix ◽  
P R Pratap ◽  
T S Novak ◽  
J C Freedman
Keyword(s):  
Membrane Potential ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Time Course ◽  
Rapid Development ◽  
Disulfonic Acid ◽  
Proton Efflux ◽  
Experimental Conditions ◽  
Human Red Blood Cells ◽  
Hydroxide Fluxes

Upon exposure of human red blood cells to hypertonic sucrose, the fluorescence of the potentiometric indicator 3,3'-dipropylthiadicarbocyanine iodide, denoted diS-C3(5), displays a biphasic time course indicating the rapid development of an inside-positive transmembrane voltage, followed by a slow DIDS (4,4'-diisothiocyano-2,2'-disulfonic acid stilbene)-sensitive decline of the voltage. In addition to monitoring membrane potential, proton (or hydroxide) fluxes were measured by a pH stat method, cell volume was monitored by light scattering, and cell electrolytes were measured directly when red cells were shrunken either with hypertonic NaCl or sucrose. Shrinkage by sucrose induced an initial proton efflux (or OH- influx) of 5.5 mu eq/g Hb.min and a Cl shift of 21-31 mu eq/g Hb in 15 min. Upon shrinkage with hypertonic NaCl, the cells are initially close to Donnan equilibrium and exhibit no detectable shift of Cl or protons. Experiments with the carbonic anhydrase inhibitor ethoxzolamide demonstrate that for red cell suspensions exposed to air and shrunken with sucrose, proton fluxes mediated by the Jacobs-Stewart cycle contribute to dissipation of the increased outward Cl concentration gradient. With maximally inhibitory concentrations of ethoxzolamide, a residual proton efflux of 2 mu eq/g Hb.min is insensitive to manipulation of the membrane potential with valinomycin, but is completely inhibited by DIDS. The ethoxzolamide-insensitive apparent proton efflux may be driven against the electrochemical gradient, and is thus consistent with HCl cotransport (or Cl/OH exchange). The data are consistent with predictions of equations describing nonideal osmotic and ionic equilibria of human red blood cells. Thus osmotic equilibration after shrinkage of human red blood cells by hypertonic sucrose occurs in two time-resolved steps: rapid equilibration of water followed by slower equilibration of chloride and protons (or hydroxide). Under our experimental conditions, about two-thirds of the osmotically induced apparent proton efflux is mediated by the Jacobs-Stewart cycle, with the remainder being consistent with mediation via DIDS-sensitive HCl cotransport (or Cl/OH exchange).

Role of chloride in potassium transport through a K-Cl cotransport system in human red blood cells

1989 ◽  
Vol 256 (5) ◽  
pp. C994-C1003 ◽  
Author(s):  
C. Brugnara ◽  
T. Van Ha ◽  
D. C. Tosteson
Keyword(s):  
Membrane Potential ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Normal Subjects ◽  
Similar Data ◽  
Human Red Blood Cells ◽  
Dense Fraction ◽  
Cotransport System ◽  
Low K

In this paper, we report experiments demonstrating the coupling of Cl and K movements in a volume-dependent K-Cl cotransport system in human red blood cells. We show that an outwardly directed Cl gradient can promote net K efflux against an inwardly directed K gradient at constant membrane potential. Red blood cell membrane potential was kept constant by using anions that are not transported through the K-Cl cotransport system but that are more permeable than Cl (NO3 and SCN). Under these conditions, when the activities of band 3 (capnophorin)-mediated anion exchange and of the carbonic anhydrase have been inhibited, it is possible to maintain a Cl gradient at constant membrane potential. Similar data were obtained in human red blood cells (least dense fraction from normal subjects and whole blood from patients with homozygous hemoglobin S disease), in rabbit red blood cells, and in low-K sheep red blood cells. These data confirm that the volume-dependent Cl-dependent K movement in these cells operates through coupled K-Cl cotransport.

scholarly journals The relation between dicarbocyanine dye fluorescence and the membrane potential of human red blood cells set at varying Donnan equilibria.

1979 ◽  
Vol 74 (2) ◽  
pp. 187-212 ◽  
Author(s):  
J C Freedman ◽  
J F Hoffman
Keyword(s):  
Membrane Potential ◽  
Fluorescence Quenching ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Red Cell Membrane ◽  
Human Red Blood Cells ◽  
Fluorescence Measurements ◽  
Constant E ◽  
Diffusion Potentials ◽  
Dye Fluorescence

The fluorescence, F, of two dicarbocyanine dyes, diS-C3(5) and diI-C3(5), depends both on the membrane potential, E, and on the intracellular pH, pHc, or human red blood cells. Compositions of isotonic media have been devised in which the equilibrium Donnan potential, E, varies at constant pHc and in which pHc varies at constant E. Dye fluorescence measurements in these suspensions yield calibrations of +1.7 % delta F/mV for diS-C3(5) and +0.6 % delta F/mV for diI-C3 (5). While pHo does not affect F of either dye, changes in pHc of 0.1 unit at constant E cause changes of F equivalent to those induced by 2--3mV. Based on these results, a method is given for estimating changes in E from dye fluorescence in experiments in which E and pHc co-vary. The relation of F to E also depends in a complex way on the type and concentration of cells and dye, and the wavelengths employed. The equilibrium calibration of dye fluorescence, when applied to diffusion potentials induced by 1 microM valinomycin, yields a value for the permeability ratio, PK.VAL/PCl, of 20 +/- 5, in agreement with previous estimates by other methods. The calibration of F is identical both for diffusion potentials and for equilibrium potentials, implying that diC-C3(5) responds to changes in voltage independently of ionic fluxes across the red cell membrane. Changes in the absorption spectra of dye in the presence of red cells in response to changes in E show that formation of nonfluorescent dimers contributes to fluorescence quenching of diS-C3(5). In contrast, only a hydrophobic interaction of dye monomers need be considered for diI-C3(5), indicating the occurrence of a simpler mechanism of fluorescence quenching.

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